The herpesviruses are ubiquitous human pathogens that are associated with conditions that include chicken pox, mononucleosis, cold sores, shingles, encephalitis, Burkitt's lymphoma and nasopharyngeal carcinoma. Infections usually persist for life in a latent state but can be reactivated with serious consequences in association with immunosuppression especially in AIDS and organ transplant patients. The complex pathways by which herpes simplex virus usurps control of cellular transcriptional machinery in lytic cycle infection presents a classical model of interactive feedback gene regulation. The levels of expression of three viral immediate-early nuclear transactivator proteins, IE175(ICP4), IE110(ICP0) and IE63(ICP27), together with the presence of the virion transactivator protein VP16 and synthesis of the latency associated modulatory RNA(LAT) are key determinants of the outcome of infection. In most cell types the presence of VP16 and the cellular Oct-1 transcription factor drives abundant synthesis of IE mRNA leading towards the lytic cycle cascade. The IE175 protein stimulates delayed-early then late gene expression, whilst simultaneously down-regulating both its own and the LAT promoters. In contrast, infection of sensory neurons in vivo results in a loss of VP16, shut-down of IE promoters and activation of LAT expression leading to establishment and maintenance of the latent state. An alternative pathway for triggering lytic cycle events, which is dependent upon IE110 function, occurs during reactivation. In this proposal we plan to continue our in vitro molecular genetic studies on the detailed mechanisms of action of the IE175 and IE110 proteins and the functional consequences of interactions between them. We will be placing particular emphasis on mapping structure/function relationships in both proteins and on identifying modular domains involved in transcriptional activation, homodimer and heterodimer formation and in promoter targeting and other protein:protein interaction events. In addition, we will continue to evaluate the mechanism of IE175 negative autoregulation by sequence specific binding to a cis-acting signal in its own cap region and to obtain an explanation for the peculiar sublocalization of IE110 in phase-dense intranuclear granules.